To understand normal brain function as well as diseases of the brain it is necessary to understand how an assembly of nerve cells can generate the processing power required for different behaviors. The vertebrate spinal cord contains a central pattern generator or CPG that can produce the rhythmic movements of locomotion in the absence of peripheral and descending inputs. CPGs serve as relatively simple model systems for understanding how neuronal assemblies produce complex functions. While the organization of the constituent interneurons in the CPG controlling swimming in non-limbed animals is known in great detail, little is known about the cellular organization of CPGs controlling walking in limbed mammals. Here we propose to identify and characterize neuronal populations in the mammalian locomotor network. We will focus on neuronal populations in segmental left-right coordination and groups of excitatory neurons that might be directly involved in rhythm generation. We will focus on these interneurons because they play key roles in CPG function. We will use in vitro preparations of prenatal and postnatal rodents for this purpose and apply a range of powerful new physiological and genetic approaches, which are adapted specifically for the proposed projects. The proposed work will provide a comprehensive anatomical, electrophysiological, pharmacological and molecular characterization of locomotor related interneurons and their connectivity. Such information will comprise 1 of the first characterizations of the roles of neuronal populations in the production of behavior by the mammalian spinal cord. Because CPG function is localized to the spinal cord and is critical for spinal control of walking, our work is of strong relevance to the ongoing effort to re-establish locomotor function in patients with spinal cord injury.